1. Field of the Disclosure
The disclosure relates to a method for analysing the effect of a test substance on biological and/or biochemical samples, particularly on samples comprising living cells. The method of the disclosure is particularly suited for the characterizing of test substances in HTS or HCS.
2. Discussion of the Background Art
High-throughput screening (HTS) and high-content screening (HCS) are methods used in the pharmaceutical and biotech industry in an effort to search for new active agents against hard-to-treat illnesses. In these methods, biochemical and/or biological samples which particularly comprise living cells, are treated with test substances such as e.g. synthetic substances, and the interaction will be analysed. “High-throughput screening” refers to the very high data throughput of the method herein (testing performed on more than 100,000 substances per day), and “high-content screening” refers to the high—particularly cellular—resolution of the signals which are to be analysed.
For performing an analysis of the effect of a test substance on a sample, particularly a cellular sample, there will first be provided a plurality of samples. These samples each comprise a known concentration of the test substance. Subsequently, measurements will be performed, particularly in high-throughput screening, wherein the samples will be e.g. illuminated by visible or non-visible light of defined wavelengths or wavelength ranges, and the radiation which is emitted by the sample, particularly due to luminescence, will be detected. In this manner, a plurality of raw data are captured per sample. Thereafter, normally, a segmentation of the obtained raw data is carried out with the aid of known image processing routines. In the process, there is performed particularly a segmentation of the cells and/or subcellular structures. Then, for the interactions to examined, an evaluation rule will be procured. From the raw data of the individual sample, a respective activity value will be obtained by use of the evaluation rule or a modelization. Said activity value describes the effect of the test substance at the present concentration. The procured evaluation rule is influenced by at least one control or model parameter. Thus, said at least one control parameter will have to be set. Since the process of setting the respective control parameters is extremely complex, this can be performed, in state-of-the-art analysis methods, only by a skilled specialist because the settings of the control parameters are normally based also on empirical values and on biological argumentation. With the aid of the so-called evaluation rule, the image data will be evaluated, and, for doses di, i=1, . . . , n, measurements mi will be obtained which quantize the interactions in the image i. In the next step, these data
      (                  d        i            ,              m        i              )        n          i      =      1      will be statistically evaluated. The statistical evaluation of the data includes e.g. the computation of the Z′-value: a statistical indicator for high-throughput screening that is an evaluation of the quality. A further statistical indicator is the dose/effect curve which represents the relation between the substance activity and the dose of the substance.
Often, the quality of the analysis depends on control parameters which have to be suitably selected. Such control parameters can be, e.g., threshold values or statistical values of the model. These will customarily be set by hand on the basis of empirical values. Further, the applicant owns a parameter scanner which is operative to search for a good setting within a finite discrete subset—which must be indicated—of all possible control parameters. The search for suitable control parameters is very bothersome.
It is an object of the disclosure, in a method for analysing the effect of a test substance on biological and/or biochemical samples, to improve the setting of the control parameters so as to accomplish an improvement of the analysis results. Particularly, it is a further object of the disclosure to provide a method of the above type wherein an automatic, preferably optimized setting of the control parameters is performed.